1. Field of the Invention
The present invention relates to mono clonal antibodies for isolating and/or identifying neural progenitor cells.
2. Related Prior Art
In contrast to many other tissues, the central nervous system has only a limited regeneration potential. Mature nerve cells which have died are not regenerated. Although neural stem cells are in fact present in the adult central nervous system (CNS), they have only a limited capacity to generate new, functionally active nerve cells after injuries.
There is for this reason great interest in the possibility of repairing the nervous system by transplanting new cells which can replace cells which have been lost through injury or disease.
At present, no suitable possibilities for remediation in particular of diseases or injuries associated with neurological deficits are available. Examples of such diseases are Parkinson's disease, Huntington's chorea, Alzheimer's disease, epilepsy, strokes or spinal cord injuries. Currently, transplantation appears to be the most promising form of therapy.
Because of the highly complicated architecture of the brain and the complex connections of the individual regions of the brain, cell replacement strategies in the nervous system make use of immature progenitor cells which must become incorpo-rated into the existing structures and do not differentiate until there.
Multipotent stem cells with the capacity of differentiating into neural cells have been found inter alia in the human central nervous system. Such neural progeni-tor cells express nestin as typical surface marker and are able to differentiate for example into neurons, oligodendrocytes and astrocytes.
Rao M S., “Multipotent and restricted precursors in the central nerv-ous system”, Anat. Rec. 257: 137 148 (1999), was able to isolate multipotent progenitor cells from adult human brain regions, including inter alia the temporal and frontal regions, the tonsils and the hippocampus. Moreover, Barami et al., “An efficient method for the culturing and generation of neurons and astrocytes from second trimester human central nervous system tissue”, Neurol. Res. 23: 321 326 (2001), showed that neural progenitor cells (NPC) from the central nervous system of human fetal tissue were CD133-positive and in addition were able to differentiate with epidermal growth factor (EGF), fibroblast growth factor and leukemia-inhibiting factor in vitro into neurons and astrocytes.
The cell surface marker CD133 was originally found on hematopoi-etic stem and progenitor cells, but more recent studies have shown that this marker is also expressed in various neural tissues and skeletal muscle tissues. For these reasons, this marker on its own is unsuitable for the purposes of distinguishing different stem or pro-genitor cells.
A great problem in the identification of neural progenitor cells is that neural progenitor cells and mesenchymal stem cells represent homogeneous populations in terms of morphology and phenotype. This is attributable in particular to the limited number of antigens investigated and identified to date.
Mesenchymal stem cells can be obtained and isolated from the bone marrow of adult humans. They are multipotent and contribute to the regeneration of bone, cartilage, tendons, muscles, adipose tissue and stroma.
Kopen et al., “Marrow stromal cells migrate throughout forebrain and cerebellum, and they differenciate into astrocytes after injection into neonatal mouse brains”, Proc. Nat. Acad. Sci. USA, 96: 10711 10716 (1999) and Brazelton et al., “From marrow to brain: expression of neuronal phenotypes in adult mice”, Science 290: 1775 1779 (2000), were in fact able to show that mesenchymal stem cells isolated from bone marrow were able to differentiate also into non-mesenchymal cells such as liver cells, neural and glial cells. In addition, it has been possible to show only recently that mesen-chymal stem cells from adult human bone marrow were able to differentiate into neural cells in vitro. Woodbury et al., “Adult rat and human bone marrow stromal cells differen-ciate into neurons”, J. Neurosci. Res. 61: 364 370 (2000), showed that in the presence of dimethyl sulfoxide (DMSO) and β-mercaptoethanol (BME) mesenchymal stem cells were able to differentiate into cells which expressed neurofilament and neuron-specific enolase.
Other research groups describe the differentiation of stromal bone marrow cells by means of epidermal growth factor and brain-derived neurotrophic factor (BDNF) into nerve cells which expressed nestin, glial fibrillary acidic protein (GFAP) and the neuron-specific nuclear protein (Neu N).
The fact that mesenchymal stem cells can also differentiate under certain conditions into nerve cells gives rise to the need to be able to distinguish neural progenitor cells from mesenchymal stem cells.
Despite the great interest, research on these neural progenitor cells has been greatly impaired by the lack of unambiguously defined markers for these cells. It is precisely the ability to identify relevant types of cells which first makes it possible to analyze the way in which the various cell populations of the central nervous system are generated.
Markers which are employed in particular for neural progenitor cells are antibodies against the protein nestin which is typically expressed by neural progenitor cells. However, this protein is also expressed by other cells such as, for example, astrocytes (see Clarke et al., “Reactive astrocytes express the embryonic intermediate neurofilament nestin”, Neuroreport 5: 1885 1888 (1994)) and muscle cells (see Sejersen and Lendahl, “Transient expression of the intermediate neurofilament nestin during skeletal muscle development”, J. Cell Sci. 106: 1291 1300 (1993)).
For these reasons, immunoreactivity with nestin is unsuitable as sin-gle criterion for identifying a particular cell as neural progenitor cell.